Numerical Density Of Synapses Research Articles

Effects of light hatching on synapse number and size in the intermediate and medial part of the hyperstriatum ventrale of the domestic chick

The effects of pre-hatch light exposure on synaptic development in the intermediate and medial part of the hyperstriatum ventrale (IMHV) of the chick brain were investigated. Quantitative electron microscopical techniques were used to determine the size and numerical density of synapses and it was seen that in light hatched chicks there was a significant increase in the density of synapses in the left IMHV but that the size of synapses in these birds was decreased. These results provide a link between synaptic development and plasticity.

Sexual differences in the numerical density of synaptic profiles of developing rat visual cortex

Axo-dendritic synaptic profiles were quantified along the whole depth of the visual cortex of 10-day-old male and female rats. In both sexes the numerical density of synaptic profiles on spine-like structures was greater than the numerical density of synapses on dendritic shafts. Females had a significantly greater numerical density of synaptic profiles on spine-like structures, than did males at a distance of 200-400 and 500-600 microns from the pia surface, which corresponds to layers II-III and IV of the cortex, respectively. A small percentage (2%-4%) of spine-like structures received two presynaptic terminals. This type of double synapses was three times more abundant in females. No sex differences were found in the numerical density of synapses on dendritic shafts in any cortical layer.

Pre‐ and postnatal development of the primary visual cortex of the common marmoset. II. Formation, remodelling, and elimination of synapses as overlapping processes

During ontogenesis changes in the numerical density of synapses are usually assumed to depend essentially on variations in the formation of synapses. Only the final adjustment to adult synapse densities is thought to include the elimination of synapses in some brain regions of certain species. Here, we focus attention on quantitative aspects of synapse elimination throughout development of area 17 of marmoset monkeys (Callithrix jacchus). Mature synapses, various precursor forms, and indicators of lysosomal degradation of synapses were quantitatively analysed by electron microscopy and morphometric methods. A total number of about 135 x 10(9) synapses was calculated for area 17 in each adult hemisphere corresponding to a volume density of 600 x 10(6) synapses/mm3. At 3 months of age, the respective values were 508 x 10(9)/area and 1,159 x 10(6)/mm3, while at birth these values were 69 x 10(9)/area and 328 x 10(6)/mm3. Consequently, at least three out of four synapses are eliminated between 3 months and adulthood. However, the real number of synapses being eliminated during development is probably much larger if the time course of lysosomal degradation is additionally taken into account. The frequency of lysosomes in presynaptic endings is highest before net-elimination of synapses occurs, i.e., between 1 and 3 months. This suggests that lysosomal degradation is not directly responsible for the majority of synapses removed during ontogenesis but apparently represents a second mechanism for synapse remodelling and elimination. Thus, it appears from this study that remodelling and elimination of synapses are quantitatively as important as their formation, and accompany synaptogenesis from its very onset onwards.

Short-term changes in the numerical density of synapses in the intermediate and medial hyperstriatum ventrale following one-trial passive avoidance training in the chick

Previous ultrastructural studies using stereological counting techniques, based on assumptions regarding shape, size, and orientation of synapses, have suggested synaptic remodeling occurred at least 24 hr after one-trial passive avoidance training in day-old chicks. The present study estimates the mean synaptic density (Nv syn) in a region of the chick forebrain known to be involved in memory formation, the intermediate and medial hyperstriatum ventrale (IMHV), 1 and 24 hr following one-trial passive avoidance training. A stereological technique, the "disector," that makes no assumptions about size, shape, and orientation of synapses was used in the synaptic analyses. The density of axospinous synapses increased by approximately 77% at 1 hr posttraining in the right IMHV of chicks (M-trained) that learned to avoid a bitter-tasting bead, compared to those (W-trained controls) that peck a water-coated bead. A measure of the postsynaptic density size, the mean projected height of synapses (H), was 57% smaller 1 hr posttraining in the right IMHV of M-trained chicks. These differences were not found at 24 hr posttraining. We suggest that structural modification of synapses may be a key part of the processes involved in short-term memory formation.

Long-term increases in the numerical density of synapses in the chick lobus parolfactorius after passive avoidance training

Passive avoidance training has been shown to cause an increase in synaptic density ( Nυ syn) in the lobus parolfactorius (LPO) of the one-day old chick 30. The present study was conducted to investigate the time-course over which this plastic change takes place. Two groups of chicks were trained to peck at either a water coated bead (Control) or a methyl-anthranilate coated bead (M-trained). M-trained chicks showed avoidance responses when offered a similar but dry bead, 30 min later. Right and left hemisphere LPOs were obtained at intervals of 1, 6, 2, 24 and 48 h after training. Synaptic counts were made using a 3-dimensional stereological probe: the ‘disector’ 27. A significantly larger mean Nυ syn (31%) was found in the left hemisphere of M-trained chicks 24 h after training, compared with Control chicks, and the difference fell to 10% at 48 h post-training. M-trained chicks also had a greater Nυ syn (17%) in the right hemisphere at 48 h post-training. The bilaterality of these findings is of particular interest, since unilateral lesions of the LPO fail to produce amnesia for the avoidance task 14. The importance of these results in the process of memory formation is discussed.

Structural asymmetry in the thalamofugal visual projections in 2-day-old chick is correlated with a hemispheric difference in synaptic density in the hyperstriatum accessorium

Differences in visual discrimination ability between the left and right eyes of chicks, which are most prominent in young males, may result from a structural asymmetry in the organization of the visual projections from the thalamus to the visual Wulst. This asymmetry in projections is no longer present by 21 days in males when the contralateral projections from the right thalamus to the left hyperstriatum have developed. Since the asymmetry of the thalamo-hyperstriatal system results in a differential input of fibres to regions of the hyperstriatum which in turn project to the hyperstriatum accessorium (HA), one of the major differences expected within this region would be an asymmetry in the numerical density of synapses ( N v,syn/ μm 3). When this was examined in the hyperstriatum of 2-day-old male chicks, the density of synapses in the right HA was found to be significantly higher (22%, P < 0.05) than in the left HA. The consequences of this asymmetry in synaptic density in the HA could be widespread and influential within the chick visual system.

Development of neurons and synapses in ochratoxin A-induced microcephalic mice: A quantitative assessment of somatosensory cortex

Ochratoxin A (a mycotoxin) is known to cause cell death in the developing brain of embryos 1–2 days after treatment. Microcephaly was observed with high frequency in mice by prenatal treatment with ochratoxin A. Using a stereological method, the numerical densities of neurons and synapses were investigated in the somatosensory cortex of 6-week-old microcephalic mice. The numerical density of neurons in ochratoxin A-treated mice represented a 39% increase compared to the control mice, but there was no difference in the numerical density of synapses. The somatosensory cortices of control mice had about 13,000 synapses per neuron, whereas ochratoxin A-treated mice had about 9,400 synapses per neuron. The deficits in synapse-to-neuron ratios seen in ochratoxin A-induced microcephalic brain seemed to result from a reduced dendritic growth.

Developmental disturbance of rat cerebral cortex following prenatal low-dose γ-irradiation: A quantitative study

Pregnant rats were exposed to a single whole-body γ-irradiation on Day 15 of gestation at a dose of 0.27, 0.48, 1.00, or 1.46 Gy. They were allowed to give birth and the offspring were killed at 6 or 12 weeks of age for microscopic and electron microscopic examinations of the cerebrum. Their body weight, brain weight, cortical thickness, and numerical densities of whole cells and synapses in somatosensory cortex were examined. Growth of the dendritic arborization of layer V pyramidal cells was also examined quantitatively with Golgi-Cox specimens. A significant dose-related reduction in brain weight was found in all irradiated groups. Neither gross malformation nor abnormality of cortical architecture was observed in the groups exposed to 0.27 Gy. A significant change was found in thickness of cortex in the groups exposed to 0.48 Gy or more. Cell packing density increased significantly in the group exposed to 1.00 Gy. Significant reduction in the number of intersections of dendrites with the zonal boundaries were found in the groups exposed to 0.27 Gy or more. There was no difference in the numerical density of synapses in layer I between the control and irradiated groups. These results suggested that doses as low as 0.27 Gy could cause a morphologically discernible change in the mammalian cerebrum.

Long-term low-protein diet reduces the number of hippocampal mossy fiber synapses

Previously we have reported a loss of the dentate granule cells and hippocampal CA3 pyramidal cells in adult animals after lengthy periods of low-protein diet. In this study we examined the effects of this cell loss upon the synaptic connections between the granule cell axons (the mossy fibers) and CA3 pyramidal cell dendrites. Three groups of five rats each were given a lowcasein (8%) diet for 6, 12, and 18 months, respectively, and the results of the ultrastructural morphometric analysis compared with similarly processed control rats kept on a control diet. The numerical density of synapses was decreased in undernourished rats and the fraction of the mossy fiber terminal membrane occupied by synaptic specializations was reduced. It can be inferred that the synaptic connectivity pattern between mossy fiber terminals and CA3 dendrites is altered due to a reduction in the number of contacts. Besides, as the synapses of low-protein-treated animals do not display any increase in the length of their active zones, evidence is not provided for the existence of morphological synaptic plasticity, contrary to what has been recently described in other experimental circumstances.

Numbers of synapses in laminae I-IV of the rat dorsal horn.

The present study determines numerical densities (NVsyn) and total numbers of synaptic discs in laminae I-IV of the rat S2 dorsal horn. Previous methods for NVsyn have the advantage of being relatively simple, but these assume that the discs are round, flat, and of uniform size. In our material, serial reconstructions indicate that these assumptions are not met. Accordingly we use a stereological method that is not as dependent on these assumptions. This method is to divide the surface density of the discs by the mean surface area of a disc (NVsyn = SVsyn/Ssyn). We refer to this as a reconstruction method because synaptic discs are reconstructed from serial sections. We also calculate numerical densities by several previously used standard methods, and the findings are similar but not identical. We find that numerical density and total synaptic numbers are smallest in lamina I, and densities and total numbers are not significantly different when lamina II is compared to laminae III and IV. Thus the intense labeling of terminals with certain compounds that characterize lamina I and II does not imply an increase in total synaptic numbers or in synaptic density. In addition there is a general increase in synaptic densities and numbers as one proceeds from lamina I to lamina IV. Another point is that the numerical density of synapses in the dorsal horn is approximately that of the cerebral cortex. These data will serve as a basis from which to judge the effects of denervations and other manipulations that purportedly change synaptic numbers.

The effects of intrauterine growth retardation on synaptogenesis and mitochondrial formation in the cerebral and cerebellar cortices of fetal sheep

In an experimental model of growth retardation which involves the reduction of placental mass in sheep, we have investigated the effects of intrauterine deprivation on synaptogenesis, synaptic ultrastructure and mitochondrial formation in the cerebral and cerebellar cortices of fetal sheep (140 days gestation). In the growth-retarded fetus, the numerical density of synapses in layer I of the visual cortex was reduced by 17% (P<0.05) compared with controls but there was no detectable difference between the two groups in the density of parallel fibre-Purkinje cell synapses in the molecular layer of the cerebellum. The length and curvature of the postsynaptic density at synapses in both regions were not affected in growth retardation but the synaptic cleft was 13% wider in the cerebellum (P<0.05) in growth retardation compared with controls. The number of mitochondrial profiles per unit area of neuropil in the visual cortex was increased by 20% (P<0.01) in growth retardation and the electron density of the inner matrix increased but the average profile area was not affected. These findings show that intrauterine growth retardation affects some aspects of synaptic development in the cerebellum and the visual cortex. The increase in the number of mitochondrial profiles in the visual cortex of growthretarded fetuses might be an attempt by the cortical neurons to compensate for the reduced efficiency of aerobic metabolism in individual mitochondria.

Alterations in synaptic structure in the paleostriatal complex of the domestic chick, Gallus domesticus, following passive avoidance training

A morphometric study was made of synapses in both left and right hemispheres of two regions of the chick paleostriatal complex, the paleostriatum augmentatum (PA) and the lobus parolfactorius (LPO), 24 h after passive avoidance training (methyl anthranilate, M-chicks), and in water-trained controls (W-chicks). The synaptic features examined were D, the mean length of the postsynaptic thickening; Nv.syn, the numerical density of synapses; Vv.syn, the volume density of the presynaptic bouton; V, the mean volume of the presynaptic bouton; Nv.ves, the numerical density of synaptic vesicles per bouton volume; ves.syn, the number of synaptic vesicles per presynaptic bouton; and K, the curvature of the synaptic contact zone. In the LPO there is a significant increase in the numerical density of synapses (Nv.syn) in both left and right hemispheres of M-compared with W-chicks (up to 59%, depending on the method of calculation used). A hemispheric asymmetry of postsynaptic thickening length (D) which is present in W-chicks (R greater than L by 10%) is reversed in M-chicks. There is no difference in the volume density of the presynaptic bouton (Vv.syn) or the mean bouton volume (V) either between W- and M-chicks, or between left and right hemispheres. Significant changes are found after avoidance training in both of the synaptic vesicle parameters measured. There is an increase of approximately 50%, both in the numerical density of synaptic vesicles (Nv.ves) and the number of vesicles per synaptic bouton (ves.syn), in the left hemisphere of M-chicks. No changes in the mean synaptic contact curvature (K) were observed after training, either of presynaptically concave, or presynaptically convex synapses, in either left or right hemispheres, nor did the percentage distribution of these different curvature classes vary greatly. In the PA there were no significant changes in D, Nv.syn or Vv.syn, either between M- and W-chicks, or between left and right hemispheres, 24 h after passive avoidance training. However, when the differences between the mean bouton volume (V) are examined, there is a significant increase in size of the boutons in the left hemisphere of M- compared to W-chicks. There is also a hemispheric asymmetry in both Nv.ves and ves.syn in W-chicks (R greater than L by approximately 15%) and this disappears on M-training. No changes in the mean synaptic contact curvature were observed after training, either of presynaptically concave, or presynaptically convex synapses, in either left or right hemispheres. However, the percentage distribution of these different curvature classes showed some variation.(ABSTRACT TRUNCATED AT 400 WORDS)

Indications for a critical period for synapse elimination in developing rat cerebral cortex cultures

It was observed in an earlier study that chronic tetrodotoxin (TTX) blockade of spontaneous bioelectric activity (SBA) in rat cerebral cortex cultures prevented the large-scale elimination of synapses which normally occurs during the fourth week in vitro. This prompted us to study whether the persisting high synapse density during long-term TTX-treatment would still return to the ‘normal’ low level after restoration of SBA. Therefore, cultures grown in TTX-supplemented medium for 5 weeks were switched to control medium for an additional week prior to fixation. Electron microscopic analysis showed that the numerical synapse density remained at a high level, thus suggesting the presence of a critical period whereafter bioelectrically controlled elimination of redundant connections no longer occurs. In contrast, the mean size of synaptic structures depended only on the functional state of the tissue at the moment of fixation, being larger in TTX-silenced cultures than in bioelectrically active ones regardless of treatment during the first 5 weeks in vitro.

Quantitative ultrastructural stereology of synapses in nucleus dorsalis after a peripheral nerve injury at birth

Utilizing recent techniques in quantitative stereology, this investigation studied the synaptology of nucleus dorsalis (Clarke's column) in 12-week-old rats whose sciatic nerves were crushed in the 1st postnatal day. Four morphometric variables were analyzed at the levels of L1 and L3 spinal cord segments: total surface area of synaptic contact zones per unit volume ( S V ), total length of synaptic contact zones per unit area ( L A ), average length of synaptic membrane ( L ), and numerical density of synapses per unit volume ( N V ). The original raw data were corrected for Holmes's effect. The results indicated that peripheral nerve crush at birth induced a transganglionic change in central sensory terminals with a loss of numerous synapses. A significant loss ( P < 0.001) of about 32% in the S V and L A and a significant loss ( P < 0.001) of about 36% in the N V were observed on the experimental side. There was no preferential loss of synapses in either segment. The mean synaptic membrane length showed no significant difference between the control and experimental sides. The control values of the four morphometric variables calculated for L3 were lower than those calculated for L1. The loss of synapses after a peripheral nerve lesion was probably due to the loss of sensory neurons and their central processes, but there were other possibilities.

Towards an improved serum-free, chemically defined medium for long-term culturing of cerebral cortex tissue

The present study describes a series of experiments which have led to a substantially improved serum-free, chemically defined medium (CDM) for long-term culturing of reaggregated fetal rat cerebral cortex tissue. A reduction of the original medium concentrations of the hormones insuline, T3 and corticosterone, on the one hand, and an enrichment of the medium with the vitamins A, C and E, the unsaturated fatty acids linoleic and linolenic acid, and biotin, L-carnitine, D(+)-galactose, glutathione (reduced) and ethanolamine, on the other hand, formed the most important chemical adjustments of the medium. With the aid of this CDM (encoded R12), the light- and electron microscopic architecture of the tissue could be kept in a good condition (superior to that seen earlier in serum-supplemented medium) up to 23 days in vitro. From that time on, the neuronal network lying between the reaggregates degenerated for the largest part, while a portion of the large neurons (probably pyramidal cells) plus some of the neuronal network within the reaggregates degenerated too. This degeneration process continued during the following weeks, but the reaggregates nevertheless retained most of their mass, so that both small and large neuronal cell bodies (visible in transparent regions at the edge of the reaggregates) remained in good condition up to at least 103 DIV. Stout, thick nerve bundles interconnecting the reaggregates, also survived up to this point. Electron microscopic evaluation of such ‘aged’ reaggregates revealed degenerating as well as healthy regions. The latter had indeed retained healthy-looking pyramidal and non-pyramidal neurons, embedded within a dense neuropil which was often traversed by myelinated axons. The numerical synapse density in such selected, healthy tissue regions reached its maximum during the sixth week in vitro, followed by a rapid decrease and a stabilization at about half the peak values. The present culture system has opened the possibility for performing controlled quantitative studies on the relationship between structure and function of cerebral cortex tissues during development and aging, on its dependence on nutrients, hormones and drugs, and on special factors synthesized by the tissue and released into the nutrient medium.

Developmental changes of synapses in the cerebellar cortex of the rat. A quantitative analysis

Synaptogenesis in the molecular layer of the vermis cortex in Wistar rats between 1 and 25 days after birth was investigated. After staining with OsO 4, the following parameters were measured: the density of synaptic profiles; the percentage of the neuropil area occupied by synapses; the mean diameter of the boutons; and the numerical density of synapses in a defined volume. The detailed topographical analysis enabled us to show the following: the first synapses appear diffusely in the molecular layer; after the 10th day a synaptic gradient is present between the depth and the surface area (this gradient can be found by the count of the synapses and by studying the spatial distribution of the synaptic area); this gradient is no longer seen at the 25th day, when the density of synapses is relatively uniform throughout the whole molecular layer.